Various feral mice are resistant to induction of erythroblastosis by Friend complex virus. In the Japanese mouse, M. m. molossinus, resistance results from inheritance of the Fv-4 resistance locus. Sensitivity to Friend complex disease was examined in these mice and in BALB/c congenics carrying Fv-4r in order to characterize Fv-4. Southern blot analysis showed that a novel ecotropic viral envelope related sequence is integrated at or near Fv-4 in resistant mice. This proviral sequence cannot be induced to produce infectious virus, but immunological methods have identified a cell surface antigen on thymocytes of Fv-4r mice related to the ecotropic envelope glycoprotein. The Fv-4r associated proviral sequence has now been molecularly cloned and biochemically characterized. The cloned fragment contains 4.0 kb of ecotropic MuLV sequences (pol, env, LTR) anc cellular flanking sequences. The nucleotide sequence of 3 kb of the viral sequences indicates that the env region resembles other ecotropic MuLVs. These data are consistent with an interference model for Fv-4 resistance in which the Fv-4r gene production binds to cell surface receptors used by ecotropic MuLVs. Other wild mice are also resistant to Friend complex disease, but lack the Fv-4 associated sequence. The resistance shown by M. spretus resembles that of Fv-2 resistant mice and results of crosses with Fv-2 sensitive and resistant inbred mice indicate that M. spretus carries the resistance allele at this locus.